Characterizing potential bacterial biocontrol agents for suppression of Rhizobium vitis, causal agent of crown gall disease in grapevines

Several bacteria were evaluated in greenhouse studies for their effectiveness at preventing formation of galls induced by Rhizobium vitis, the Gram negative bacterium that causes crown gall disease of grapevines. Three endophytic bacteria isolated from Malus domestica yielded promising results and were selected for further examination: Pseudomonas fluorescens isolate 1100-6, Bacillus subtilis isolate EN63-1, and Bacillus species isolate EN71-1. In planta experiments were initially performed on Nicotiana glauca to facilitate measurement of gall development. The effect of the interval between inoculation of N. glauca with the biocontrol agent and exposure to R. vitis was evaluated. All three bacteria significantly reduced gall size when applied 25 or 86 days before inoculation with R. vitis, and reduced the population of R. vitis relative to control plants. P. fluorescens '1100-6' also reduced the percentage of R. vitis inoculations that yielded galls. P. fluorescens '1100-6' was genetically tagged to allow exploration of its localization in grapevines. In growth chamber studies, P. fluorescens '1100-6' was found to survive in the rhizoplane of grapevines for 6 months. Microscopic examination showed that the genetically modified P. fluorescens '1100-6' predominantly occupied xylem and pith tissues, and occasionally persisted on external surfaces of grapevines.